A Simple and Accurate Reverse Phase HPLC-UV Method for Determination of Gossypol and Its Degradation Products

Gossypol is a toxic polyphenolic compound present in cotton plant including its seed. Microbial fermentation using Pleurotus flabellatus M-1 for 72 h degraded gossypol to 76.6% in minimal medium. A simple, precise and accurate method for gossypol determination was developed and validated. A high performance liquid chromatographic method (HPLC) consisting of an isocratic mobile phase of acetonitrile: water (80: 20, v/v) with 0.1% phosphoric acid was used for the resolution of gossypol on RP-C₁₈column (4.6 mm × 250 mm, 5μm particle), at a flow rate of 1.0 ml min⁻¹ and UV detection at 254 nm. Under these conditions, the gossypol and its degradation products were separated in 12.5 and 8.3 min retention time, respectively. The modified method was accurate, precise, and reproducible with linearity (R²=0.999, n=5, P<0.05), limit of detection (LOD=0.2 μg ml⁻¹) and limit of quantification (LOQ=0.5 μg ml⁻¹). Absolute recoveries were greater than 97.6% with an average RSD of 0.75%. This method could probably be scaled up for researchers for accurate gossypol quantification as well as for identifying its degradation products.